Small molecule-mediated allosteric activation of the base excision repair enzyme 8-oxoguanine DNA glycosylase and its impact on mitochondrial function

Abstract

Abstract8-Oxoguanine DNA glycosylase (OGG1) initiates base excision repair of the oxidative DNA damage product 8-oxoguanine. OGG1 is bifunctional; catalyzing glycosyl bond cleavage, followed by phosphodiester backbone incision via a β-elimination apurinic lyase reaction. The product from the glycosylase reaction, 8-oxoguanine, and its analogues, 8-bromoguanine and 8-aminoguanine, trigger the rate-limiting AP lyase reaction. The precise activation mechanism remains unclear. The product-assisted catalysis hypothesis suggests that 8-oxoguanine and analogues bind at the product recognition (PR) pocket to enhance strand cleavage as catalytic bases. Alternatively, they may allosterically activate OGG1 by binding outside of the PR pocket to induce an active-site conformational change to accelerate apurinic lyase. Herein, steady-state kinetic analyses demonstrated random binding of substrate and activator. 9-Deazaguanine, which can’t function as a substrate-competent base, activated OGG1, albeit with a lower Emax value than 8-bromoguanine and 8-aminoguanine. Random compound screening identified small molecules with Emax values similar to 8-bromoguanine. Paraquat-induced mitochondrial dysfunction was attenuated by several small molecule OGG1 activators; benefits included enhanced mitochondrial membrane and DNA integrity, less cytochrome c translocation, ATP preservation, and mitochondrial membrane dynamics. Our results support an allosteric mechanism of OGG1 and not product-assisted catalysis. OGG1 small molecule activators may improve mitochondrial function in oxidative stress-related diseases.

The evolution of small molecule enzyme activators

“…These compounds were also identified to reduce cytochrome c concentration (reduction of mean nuclear intensity by 550 for both 30 & 31 at 30 μM), which is associated with mitochondrial membrane integrity and DNA damage. 213 It has further been reported that mitochondrial DNA levels in mouse embryonic fibroblasts were recovered at a faster rate when treated with 31 than alone when cells are subjected to hydrogen peroxide. 221 To identify the mechanism in which 32 activates OGG1 in vitro , DNA repair studies and cocrystal structure analysis were undertaken.…”

“…The bifunctional enzyme 8-oxoguanine DNA glycosylase (OGG1) catalyzes both glycosylase and abasic (AP) lyase reactions, with the latter being the rate-limiting step. 211–213 Mutagenic experiments have identified key residues in the active site that are crucial for activity, resulting in multiple postulated mechanisms. 214 Firstly, Lys 249 acts as a catalytic nucleophile in the glycosylase reaction, this allows for a covalent enzyme–DNA interaction which can undergo rearrangement.…”

“…20) have been identified as activators of OGG1 in a fluorescent kinetic assay with EC 50 's of 4.1 μM, 5.1 μM, 4.6 μM, and 5.0 μM respectively, for 28-31, and an AC 50 of 0.78 μM for 32. 213,220,221 These compounds were also assessed for their ability to reduce A549 cell injury caused under oxidative challenge, accumulation of 8-oxo-G, and associated induced cytotoxicity. This experiment identified compounds 29 and 31 can reduce cell injury by oxidative damage (by 200% and 650%, respectively).…”

Complex Roles of NEIL1 and OGG1: Insights Gained from Murine Knockouts and Human Polymorphic Variants

“…In this regard, the initial report of an agonist of OGG1 identified 8-bromoGua, which showed significant stimulation of the OGG1 lyase activity [74,184]. Additional studies have reported the identification of novel OGG1 agonists [163], with recent follow-up studies determining the underlying mechanisms of the AP lyase catalytic function and the benefits of enhanced mitochondrial repair [185]. Recently, another potential activator of OGG1 has been described in which the AP lyase activity of OGG1 was stimulated via small-molecule screening [186].…”

8-Oxoguanine DNA Glycosylase 1 Upregulation as a Risk Factor for Obesity and Colorectal Cancer

“…ROS can damage DNA in different pathways. However, 8-oxoguanine (8-oxoG) stands out as one of the most common DNA lesions observed as a result of exposure to ROS products [ 6 ]. 8-OxoG is a tautomer that can result in a mismatched pairing with adenine [ 7 ].…”